title: An insertion peptide in yeast glycyl-tRNA synthetase facilitates both productive docking and catalysis of cognate tRNAs.
abstract: The yeast Saccharomyces cerevisiae possesses two distinct glycyl-tRNA synthetase (GlyRS) genes: GRS1 and GRS2. GRS1 is dually functional, encoding both cytoplasmic and mitochondrial activities, while GRS2 is dysfunctional and not required for growth. The protein products of these two genes, GlyRS1 and GlyRS2, are much alike but are distinguished by an insertion peptide of GlyRS1, which is absent from GlyRS2 and other eukaryotic homologues. We show that deletion or mutation of the insertion peptide modestly impaired the enzyme's catalytic efficiency in vitro (with a 2- to 3-fold increase in Km and a 5- to 8-fold decrease in kcat). Consistently, GRS2 can be conveniently converted to a functional gene via codon optimization, and the insertion peptide is dispensable for protein stability and the rescue activity of GRS1 at 30°C in vivo. A phylogenetic analysis further showed that GRS1 and GRS2 are paralogues that arose from a gene duplication event relatively recently, with GRS1 being the predecessor. These results indicate that GlyRS2 is an active enzyme essentially resembling the insertion peptide-deleted form of GlyRS1. Our study suggests that the insertion peptide represents a novel auxiliary domain, which facilitates both productive docking and catalysis of cognate tRNAs.
authors.affiliation: Department of Life Sciences, National Central University, Jungli, Taiwan.
authors.forename: Yi-Hua
authors.initials: YH
authors.lastname: Wu
authors.forename: Chia-Pei
authors.initials: CP
authors.lastname: Chang
authors.forename: Chin-I
authors.initials: CI
authors.lastname: Chien
authors.forename: Yi-Kuan
authors.initials: YK
authors.lastname: Tseng
authors.forename: Chien-Chia
authors.initials: CC
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D003062:Codon; D020440:Gene Duplication; D005800:Genes, Fungal; D006032:Glycine-tRNA Ligase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation; D055550:Protein Stability; D012343:RNA, Transfer / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017384:Sequence Deletion; D013696:Temperature
chemical_list: D003062:Codon; D029701:Saccharomyces cerevisiae Proteins; D012343:RNA, Transfer; C478769:GRS1 protein, S cerevisiae; C583888:GRS2 protein, S cerevisiae; D006032:Glycine-tRNA Ligase